Human granulocytic anaplasmosis is the second most common tick-borne disease in the United States. The causative agent, Anaplasma phagocytophilum, is an obligate intracellular pathogen that resides in 2 different locations during its life cycle -- mammalian neutrophils and Ixodes scapularis salivary glands. This project investigates whether tick genes that are up- or down-regulated in the presence of A. phagocytophilum play important roles in infection. These studies build upon our efforts showing that A. phagocytophilum alters host gene expression and uses specific molecules for survival. As examples, in mammals Anaplasma requires fucosylation of host proteins for infection, and represses gp91phox in neutrophils to prevent the respiratory burst. In ticks Anaplasma induces I. scapularis salivary protein (Salp) 16 to colonize the vector by mechanisms that are currently not known. Our preliminary data also show that A. phagocytophilum infection of I. scapularis induces a tick fucosyltransferase (IsFucoT), and decreases the expression of I. scapularis salp10 and salp17. We now hypothesize that salp16 and IsFucoT - 2 Anaplasma-induced tick genes - are important for A. phagocytophilum infection of I. scapularis and will elucidate how this occurs. This is based, in part, on our preliminary data showing that RNA interference-mediated knockdown of IsFucoT decreases A. phagocytophilum infection of I. scapularis. We will also examine whether Salp16 is fucosylated, thereby providing an intimate linkage between these 2 Anaplasma-induced tick genes. We also hypothesize that salp10 and salp17 - two tick genes that are repressed during A. phagocytophilum infection of I. scapularis - are also crucial in the Anaplasma life cycle. We will therefore characterize salp10 and salp17, and use a lentiviral-based system to introduce these genes into ticks -- the first "knock-in" studies to generate ticks that constitutively express a specific gene. We postulate that A. phagocytophilum survival will be altered in ticks when salp10 and salp17 are not normally down-regulated during infection. These studies will provide an understanding of how tick genes that are up- or down-regulated in the presence of A. phagocytophilum are important for infection of I. scapularis, delineate similarities and differences between infection in the arthropod vector and mammalian host, and lead to new strategies to interfere with the Anaplasma life cycle. PUBLIC HEALTH RELEVANCE: Human granulocytic anaplasmosis (HGA), previously known as human granulocytic ehrlichiosis, is the second most common tick-borne illness in North America. The causative agent is Anaplasma phagocytophilum, an obligate intracellular pathogen that persists within neutrophils in the mammalian host, and also in cells within Ixodes scapularis ticks. The goal of this proposal is to develop a better understanding of the relationship between A. phagocytophilum and its tick vector, I. scapularis. A. phagocytophilum alters the expression of several mammalian genes during infection of neutrophils, and this pathogen also influences the expression of specific I. scapularis genes. By understanding these A. phagocytophilum-tick interactions in greater detail, we will develop new ways to interfere with the A. phagocytophilum life cycle. These studies should lead to a greater understanding of A. phagocytophilum pathogenesis and may suggest new ways to prevent infection. It is also hopeful that these paradigms would also be applicable to other vector-borne pathogens of medical importance.